Protein translation typically begins with the recruitment of the 43S ribosomal complex to the 5′ cap of mRNAs by a cap-binding complex. However, some transcripts are translated in a cap-independent manner through poorly understood mechanisms. For most cellular mRNAs, the first step of mRNA translation involves recognition of the 5′ 7-methylguanosine (m7G) cap by eukaryotic initiation factor 4E (eIF4E), which is a subunit of the heterotrimeric eIF4F complex. 5′ cap-bound eIF4F then recruits the small (40S) ribosomal subunit associated with various translation initiation factors, enabling efficient translation of eukaryotic mRNAs.
However, some mRNAs are translated in a cap-independent manner. These capped mRNAs do not require eIF4E and are translated under basal cellular conditions as well as conditions where eIF4E activity is compromised, such as cellular stress states, viral infection, and in diseases such as cancer (Stoneley et al., “Cellular Internal Ribosome Entry Segments: Structures, Trans-Acting Factors and Regulation Of Gene Expression,” Oncogene 23:3200-3207 (2004)). Although viral mRNAs can exhibit cap-independent translation due to the presence of highly structured internal ribosome entry site (“IRES”) motifs in the 5′ UTR, correspondingly complex structures are rarely found in eukaryotic mRNAs undergoing cap-independent translation (Stoneley et al., “Cellular Internal Ribosome Entry Segments: Structures, Trans-Acting Factors and Regulation Of Gene Expression,” Oncogene 23:3200-3207 (2004)). Thus, the mechanism of cap-independent translation in cellular mRNAs remains poorly understood.
A feature of many eukaryotic mRNAs is N6-methyladenosine (“m6A”), a reversible base modification seen in the 3′ UTR coding sequence, and 5′ UTR (Dominissini et al., “Topology of the Human and Mouse m6A RNA Methylomes Revealed by m6A-Seq.,” Nature 485:201-206 (2012); Meyer et al., “Comprehensive Analysis of mRNA Methylation Reveals Enrichment in 3′ UTRs and Near Stop Codons,” Cell 149:1635-1646 (2012). Although the function of m6A in the coding sequence and 3′ UTRs has been explored (Wang et al., “N6-Methyladenosine-Dependent Regulation of Messenger RNA Stability,” Nature 505:117-120 (2014a); Wang et al., “N(6)-Methyladenosine Modulates Messenger RNA Translation Efficiency,” Cell 161:1388-1399 (2015); Wang et al., “N(6)-Methyladenosine Modification Destabilizes Developmental Regulators in Embryonic Stem Cells,” Nat. Cell Biol. 16:191-198 (2014b)), the function of m6A in 5′ UTRs remains unknown.
The present invention is directed to overcoming these and other deficiencies in the art.